Study On Rapid Propagation Of Swertia Pseudochinensis Hara And Physiological And Biochemical Characteristics Of Callus

Swertia pseudochinensis Hara is an important Mongolian medicine belonging to Swertiaof Gentianageae,which has the effect of clearing away damp-heat.At present,the medicinal materials are mainly derived from the wild.Due to the drastic reduction of wild resources,it has been unable to meet the market demand.Plant tissue culture technology has been widely used in the rapid propagation of plants,and provides a practical and feasible way for the sustainable use of traditional Chinese medicine resources.Therefore,the rapid propagation of tissue culture and related technologies in S.pseudochinensis can effectively solve the shortage of medicinal materials.Environmental variables are important factors affecting the distribution of S.pseudochinensis.Analysis of the key environmental factors in its habitat can help the cultivation and introduction of S.pseudochinensis.According to the random forest regression analysis of the importance of variables,the most important environmental variable affecting the distribution of S.pseudochinensis is the annual precipitation,followed by the annual average temperature,indicating that humidity and temperature affect the physiological characteristics of S.pseudochinensis.Disinfection is a prerequisite for the establishment of rapid propagation system of plant tissue culture seedlings.For the establishment of a rapid propagation system using mature seeds as explants,the disinfecting conditions are as follows: 70% of 84 disinfectants are used to disinfection for 3 minutes.Then MS is used as the basic medium at a temperature of(25±1)?.When the light intensity is 1500 lx and the illumination time is 12 h/d,cultured for 30 days,sterile seedlings could be obtained.Through the screening of the dedifferentiation and re-differentiation conditions of S.pseudochinensis,the final results show that the induction effects of different explants on callus are significantly different.The induction effect of tender stems is better,and the recovery time is early and the induction rate is high.Under light illumination,the optimum induction medium for tender stems is MS+2.0 mg/L 2,4-D +0.3mg/L ZT +1.0 mg/L6-BA;under dark conditions,the optimal induction medium is MS+1.0 mg/ L2,4-D+0.5mg/L6-BA+0.3mg/L ZT;optimal proliferation medium is MS+1.0mg/L 6-BA+0.3mg/LNAA;optimal differentiation medium is MS+0.1mg/LNAA+1.0 mg/L6-BA.By comparing the physiological and biochemical characteristics of different callus in the process of tissue culture,I find that the protein content and soluble sugar content of embryogenic callus are 41.11mg/g and 15.91mg/g,respectively,which are significantly higher than those of non-embryonic callus.The amount of CAT contained in the tissue is significantly higher than that of the non-embryogenic callus;the SOD content is lower than that of the non-embryonic callus but do not reach a significant level,and the POD content is significantly lower than that of the non-embryonic callus.It shows that the physiological activity andmetabolism ofembryogenic callus are more active than the non-embryogenic callus.Through the study of physiological and biochemical changes during the somatic embryogenesis of S.pseudochinensis,it is found that the change of soluble sugar content decreases first,then increases,then decreases;the change of soluble protein content increases sharply from 28 to 35 days,and other times are relatively stable;SOD activity and POD activity show an overall upward trend;PPO activity decreases first and then increases.It indicates that the physiological and biochemical indexes of somatic embryos change correspondingly in the content and activity of callus at different stages,which lay an important theoretical foundation for the later differentiation of callus and the optimization of culture conditions for tissue culture.Through the preliminary determination of the main medicinal components in the callus and regenerated plants of S.pseudochinensis,the results show that the content of tartarin in the original plant is 0.028%,the content of swertiamarin is 0.68%;The contents in the callus and tissue culture seedlings are lower,indicating that there are differences in the efficacy components in different materials,and the causes of the differences need to be further studied.In summary,by studying the physiological and biochemical changes in the tissue culture of S.pseudochinensis,it lays a scientific foundation for the next step in the establishment of tissue culture and rapid propagation.